Revisiting the Extended X-ray Absorption Fine Structure Fitting Procedure through a Machine Learning-Based Approach.

A novel approach for the analysis of extended X-ray absorption fine structure (EXAFS) spectra is developed exploiting an inverse machine learning-based algorithm. Through this approach, it is possible to explore and account for, in a precise way, the nonlinear geometry dependence of the photoelectron backscattering phases and amplitudes of single and multiple scattering paths. In addition, the determined parameters are directly related to the 3D atomic structure, without the need to use complex parametrization as in the classical fitting approach. The applicability of the approach, its potential and the advantages over the classical fit were demonstrated by fitting the EXAFS data of two molecular systems, namely, the KAu (CN)2 and the [RuCl2(CO)3]2 complexes.

TRANSPARENT TESTA 16 and 15 act through different mechanisms to control proanthocyanidin accumulation in Arabidopsis testa.

Flavonoids are secondary metabolites that fulfil a multitude of functions during the plant life cycle. In Arabidopsis proanthocyanidins (PAs) are flavonoids that specifically accumulate in the innermost integuments of the seed testa (i.e. endothelium), as well as in the chalaza and micropyle areas, and play a vital role in protecting the embryo against various biotic and abiotic stresses. PAs accumulation in the endothelium requires the activity of the MADS box transcription factor TRANSPARENT TESTA (TT) 16 (ARABIDOPSIS B-SISTER/AGAMOUS-LIKE 32) and the UDP-glycosyltransferase TT15 (UGT80B1). Interestingly tt16 and tt15 mutants display a very similar flavonoid profiles and patterns of PA accumulation. By using a combination of genetic, molecular, biochemical, and histochemical methods, we showed that both TT16 and TT15 act upstream the PA biosynthetic pathway, but through two distinct genetic routes. We also demonstrated that the activity of TT16 in regulating cell fate determination and PA accumulation in the endothelium is required in the chalaza prior to the globular stage of embryo development. Finally this study provides new insight showing that TT16 and TT15 functions extend beyond PA biosynthesis in the inner integuments of the Arabidopsis seed coat.

Pre-adapting parasitic phages to a pathogen leads to increased pathogen clearance and lowered resistance evolution with Pseudomonas aeruginosa cystic fibrosis bacterial isolates.

Recent years have seen renewed interest in phage therapy--the use of viruses to specifically kill disease-causing bacteria--because of the alarming rise in antibiotic resistance. However, a major limitation of phage therapy is the ease at with bacteria can evolve resistance to phages. Here, we determined whether in vitro experimental coevolution can increase the efficiency of phage therapy by limiting the resistance evolution of intermittent and chronic cystic fibrosis Pseudomonas aeruginosa lung isolates to four different phages. We first pre-adapted all phage strains against all bacterial strains and then compared the efficacy of pre-adapted and nonadapted phages against ancestral bacterial strains. We found that evolved phages were more efficient in reducing bacterial densities than ancestral phages. This was primarily because only 50% of bacterial strains were able to evolve resistance to evolved phages, whereas all bacteria were able to evolve some level of resistance to ancestral phages. Although the rate of resistance evolution did not differ between intermittent and chronic isolates, it incurred a relatively higher growth cost for chronic isolates when measured in the absence of phages. This is likely to explain why evolved phages were more effective in reducing the densities of chronic isolates. Our data show that pathogen genotypes respond differently to phage pre-adaptation, and as a result, phage therapies might need to be individually adjusted for different patients.

Evolution of the active species of homogeneous Ru hydrodeoxygenation catalysts in ionic liquids.

This work establishes structure-property relationships in Ru-based catalytic systems for selective hydrodeoxygenation of ketones to alkenes by combining extensive catalytic testing, in situ X-ray absorption spectroscopy (XAS) under high pressures and temperatures and ex situ XAS structural characterization supported by density functional theory (DFT) calculations. Catalytic tests revealed the difference in hydrogenation selectivity for ketones (exemplified by acetone) or alkenes (exemplified by propene) upon changing the reaction conditions, more specifically in the presence of CO during a pretreatment step. XAS data demonstrated the evolution of the local ruthenium structure with different amounts of Cl/Br and CO ligands. In addition, in the absence of CO, the catalyst was reduced to Ru0, and this was associated with a significant decrease of the selectivity for ketone hydrogenation. For the Ru-bromide carbonyl complex, selectivity towards acetone hydrogenation over propene hydrogenation was explained on the basis of different relative energies of the first intermediate states of each reaction. These results give a complete understanding of the evolution of the Ru species, used for the catalytic valorization of biobased polyols to olefins in ionic liquids, identifying the undesired deactivation routes as well as possibilities for reactivation.

Photopatterning of fluorescent host-guest carriers through pore activation of metal-organic framework single crystals.

Encoded fluorescent particles are fabricated through the selective uptake of dyes in photopatterned metal-organic framework single crystals. The concept is based on spatially controlled photochemical cleavage of pore-blocking pendant groups. Because of the crystalline and porous nature of the host, this approach enables guest uptake that is tunable and can be triggered though controlled irradiation.

Study of the active site residues of a glycoside hydrolase family 8 xylanase.

Site-directed mutagenesis and a comparative characterisation of the kinetic parameters, pH dependency of activity and thermal stability of mutant and wild-type enzymes have been used in association with crystallographic analysis to delineate the functions of several active site residues in a novel glycoside hydrolase family 8 xylanase. Each of the residues investigated plays an essential role in this enzyme: E78 as the general acid, D281 as the general base and in orientating the nucleophilic water molecule, Y203 in maintaining the position of the nucleophilic water molecule and in structural integrity and D144 in sugar ring distortion and transition state stabilization. Interestingly, although crystal structure analyses and the pH-activity profiles clearly identify the functions of E78 and D281, substitution of these residues with their amide derivatives results in only a 250-fold and 700-fold reduction in their apparent k(cat) values, respectively. This, in addition to the observation that the proposed general base is not conserved in all glycoside hydrolase family 8 enzymes, indicates that the mechanistic architecture in this family of inverting enzymes is more complex than is conventionally believed and points to a diversity in the identity of the mechanistically important residues as well as in the arrangement of the intricate microenvironment of the active site among members of this family.

Towards metal-organic framework based field effect chemical sensors: UiO-66-NH2 for nerve agent detection.

We present a highly sensitive gas detection approach for the infamous 'nerve agent' group of alkyl phosphonate compounds. Signal transduction is achieved by monitoring the work function shift of metal-organic framework UiO-66-NH2 coated electrodes upon exposure to ppb-level concentrations of a target simulant. Using the Kelvin probe technique, we demonstrate the potential of electrically insulating MOFs for integration in field effect devices such as ChemFETs: a three orders of magnitude improvement over previous work function-based detection of nerve agent simulants. Moreover, the signal is fully reversible both in dry and humid conditions, down to low ppb concentrations. Comprehensive investigation of the interactions that lead towards this high sensitivity points towards a series of confined interactions between the analyte and the pore interior of UiO-66-NH2.

Access to bacteriophage therapy: discouraging experiences from the human cell and tissue legal framework.

Cultures of human epithelial cells (keratinocytes) are used as an additional surgical tool to treat critically burnt patients. Initially, the production environment of keratinocyte grafts was regulated exclusively by national regulations. In 2004, the European Tissues and Cells Directive 2004/23/EC (transposed into Belgian Law) imposed requirements that resulted in increased production costs and no significant increase in quality and/or safety. In 2007, Europe published Regulation (EC) No. 1394/2007 on Advanced Therapy Medicinal Products. Overnight, cultured keratinocytes became (arguably) 'Advanced' Therapy Medicinal Products to be produced as human medicinal products. The practical impact of these amendments was (and still is) considerable. A similar development appears imminent in bacteriophage therapy. Bacteriophages are bacterial viruses that can be used for tackling the problem of bacterial resistance development to antibiotics. Therapeutic natural bacteriophages have been in clinical use for almost 100 years. Regulators today are framing the (re-)introduction of (natural) bacteriophage therapy into 'modern western' medicine as biological medicinal products, also subject to stringent regulatory medicinal products requirements. In this paper, we look back on a century of bacteriophage therapy to make the case that therapeutic natural bacteriophages should not be classified under the medicinal product regulatory frames as they exist today. It is our call to authorities to not repeat the mistake of the past.

Studies of decitabine with allogeneic progenitor cell transplantation.

The aim was to determine the efficacy and safety of decitabine in the settings of relapse post-allogeneic progenitor cell transplantation or as part of the conditioning regimen. Three patients (two AML, one ALL) received single agent decitabine 1000 mg/m2 total dose) for treatment of relapse post-transplant (group 1). Median age was 32 years. Median time to relapse was 7 months. In another study four patients (three CML in an accelerated phase, one AMML) received decitabine 400 mg/m2, with busulfan 12 mg/kg and cyclophosphamide 100 mg/kg as conditioning for allogeneic stem cell transplantation (group 2). Median age was 42 years; median time to transplant was 5 months. All patients received at least 4 x 10(6) CD34+ cells from their HLA compatible donors. All patients in group 1 achieved complete remissions after decitabine therapy. The median time to neutrophil and platelet recovery were 24 and 23 days, respectively. Two patients required reinfusion of donor cells because of delayed engraftment. One patient remains alive and in remission 160 days post-decitabine therapy. Two patients in group 2 engrafted on days 23 and 25. Two patients required reinfusion of stem cells because of lack of neutrophil recovery by day 21. Two patients achieved complete cytogenetic and hematologic remission. Three patients are alive at 167,129, and 109 days post-transplant. One patient died of progressive Pseudomonas cellulitis 54 days post-initial infusion. Decitabine therapy is well tolerated in the setting of allogeneic stem cell transplantation, initial results in patients relapsing after transplant are encouraging and warrant further studies. The causes of delayed engraftment after single agent or combination therapy need to be better explored. The existence of active metabolites of decitabine which may still be present in the blood at the time of stem cell infusions, and/or insufficient immunosuppression of the preparative regimen are being explored as possible explanations for this phenomenon.

Results of decitabine therapy in the accelerated and blastic phases of chronic myelogenous leukemia.

The aim of the study was to evaluate the activity of decitabine, a hypomethylating agent, in the treatment of patients with chronic myelogenous leukemia (CML) in transformation. Thirty-seven patients with CML in blastic (20 patients) or accelerated phases (17 patients) were treated. Their median age was 52 years; 36 had Philadelphia chromosome-positive disease. Decitabine was given at 100 mg/m2 over 6 h every 12 h x 10 doses (1000 mg/m2) to 13 patients, and at 75 mg/m2 over 6 h every 12 h x 10 doses (750 mg/m2) to 24 patients. In blastic phase, two patients (10%) achieved a complete hematologic response (one with Ph suppression), and three (15%) had a hematologic improvement (marrow CR, platelets <100 x 10[3]/microl), for an overall response rate of 25%. In accelerated phase, six patients (35%) returned to a second chronic phase (two with Ph suppression), one (6%) had a hematologic improvement, and two (12%) had a partial hematologic response, for an overall response rate of 53%. Prolonged myelosuppression was the most significant side-effect. The median time to recovery of granulocytes above 500/microl was 48 days, and to recovery of platelets above 30 x 10(3)/microl, 31 days. Febrile episodes occurred in 25 patients (68%) including documented infections in 17 patients (46%). Decitabine has promising activity in CML. The most significant side-effect is prolonged myelosuppression. Decitabine may show activity in other myeloid disorders such as acute myeloid leukemia and myelodysplastic syndrome, as well as in other hematologic malignancies, alone or with other drug combinations. Its value in the context of stem cell support should also be investigated.

Antitumour activity and retinotoxicity of ethyldeshydroxy-sparsomycin in mice.

The colony formation in agar of human tumour xenografts was used as a test system to study the cytostatic activity of ethyldeshydroxy-sparsomycin (EdSm) at the cellular level. EdSm was additionally studied in vivo in human tumour xenografts and murine tumour models. EdSm showed a clear dose-response effect in vitro. At continuous exposure with 0.01 micrograms/ml, 2 out of 11 of the tumours responded (a gastric and a small cell lung carcinoma). At 0.1 mu/ml EdSm, the tumour response was 5/11 tumours and at 1 microgram/ml the compound was active in all tumours. The maximal tolerable doses of EdSm in vivo have been determined in non-tumour bearing CDF1 mice. In the intraperitoneally (i.p.) given multiple dose schedules the respective LD10 doses indicated that the tolerable cumulative dose increases when lower doses are given more frequently. This also enhances the antitumour activity in L1210 leukaemia to 172% T/C. On the other hand, continuous infusion strongly diminished the tolerable dose as well as the antitumour activity. EdSm was also active against i.p. inoculated P388 leukaemia (150% T/C), B16 melanoma (156% T/C), and RC carcinoma (197% T/C), and the subcutaneously (s.c.) inoculated L1210 (139% T/C) and RC (138% T/C). Absence of tumour responses was found in the following s.c. implanted murine tumours: M5076 sarcoma, osteosarcomas C22LR and CP369, and the LL carcinoma, as well as in the human tumour xenografts: LXFG 529, a non-small cell lung carcinoma; GXF 251, a gastric carcinoma; and FMa, an ovary carcinoma. Possible long-range retinotoxic effects of EdSm were investigated in tumour-bearing mice, cured after surviving treatment with LD50 doses of EdSm, by assaying the protein biosynthetic capacity of the retinal by assaying the ocular rhodopsin and opsin levels as parameters. In none of these cases could a significant reduction in either opsin or rhodopsin levels be measured and no changes were seen histologically.

Serum elimination half-life of tamoxifen and its metabolites in patients with advanced breast cancer.

In breast cancer patients discontinuing chronic tamoxifen therapy, the serum elimination of metabolites X, Y and E paralleled that of tamoxifen, whereas that of metabolite Z did not. The serum elimination of tamoxifen and metabolites X and B was increased by amino-glutethimide treatment, whereas that of metabolites Z, Y, and E was not.

Serum and urine levels of tamoxifen and its metabolites in patients with advanced breast cancer after a loading dose and at steady-state levels.

PURPOSE: To compare serum and urine levels of tamoxifen and metabolites after a loading dose and at the steady state. METHODS: A loading dose of 160 mg of tamoxifen was given to 14 patients with advanced breast cancer. Thereafter a regular daily dose of 30 mg of tamoxifen was given. Serum and urine levels of tamoxifen and metabolites were measured by high-performance liquid chromatography and compared with levels determined in 31 patients with advanced breast cancer at the steady state at a daily dose of 30 mg of tamoxifen. RESULTS: Serum and urine levels (24-h values) of tamoxifen and metabolites were lower (P < 0.05) after a loading dose than at the steady state. The difference was most pronounced for the metabolites, whereas the tamoxifen loading-dose level was near the steady state. CONCLUSION: Tamoxifen steady state can be reached in 1-2 days by the administration of a loading dose of 160 mg of tamoxifen for 2 days. Tamoxifen metabolite steady-state levels are reached regularly after 4 or more weeks during application of a loading dose. Very little tamoxifen or metabolites are excreted into the urine.

Impact of Pd-mordenite pretreatment on the heterogeneity of Heck catalysis.

In Heck reactions with tributylamine as the base and in toluene, Pd(NH3)4(2+)-mordenite (0.4 wt% Pd) and Pd0-mordenite (0.4 and 4 wt% Pd) are not only active and selective, but also truly heterogeneous catalysts, while oxidized PdII species in an all-oxygen environment, i.e. ionic Pd2+ or PdO, are prone to leaching.

In vitro study of the photocytotoxicity of some hypericin analogs on different cell lines.

In the present study, hypericin analogs with an increased hydrophilic character were synthesized. As chemical modifications alter the lipophilicity/hydrophilicity balance together with the photophysical/chemical background of the molecule the influence of these structural changes on the cellular uptake, retention and subcellular localization in HeLa cells was investigated. Besides, their photocytotoxic effects using three cell lines (HeLa, MCF-7, A431), as well as their plasma protein binding were also assessed. To assess the relative hydrophilic/lipophilic character of hypericin and analogs their retention times were determined on a reversed phase high performance liquid chromatography (C-18) column. The retention time of all the hypericin analogs was < 46 min, except for dibenzyltetramethylhypericin (118 min), while the retention time of hypericin was > 200 min (solvent system: methanol/citrate buffer 30 mM pH 7; 70/30). Hypericin, hexa-, penta- and dibenzyltetramethylhypericin displayed a potent antiproliferative effect at the nanomolar range after photosensitization (3.6 J/cm2). On the contrary, photoactivated tetrasulfonhypericin and fringelite D had no antiproliferative effect on the three cell lines, whereas hypericin polyethylene glycol showed only an intermediate cytotoxic effect on A431 cells. In dark conditions no antiproliferative effect was observed for any photosensitizer. The antiproliferative photo-effect correlated well with the intracellular accumulation as measured using HeLa cells. In general, the photocytotoxic hypericin analogs concentrated to a large extent, while the noncytotoxic compounds were not taken up by the HeLa cells. Furthermore, confocal laser microscopy revealed that all photosensitizers mainly concentrated in the perinuclear region, probably corresponding with Golgi apparatus and the endoplasmic reticulum, except for tetrasulfonhypericin which located at the plasma membrane. In addition, the plasma protein binding studies illustrated that hypericin bind extensively to the low-density lipoproteins, while the other hypericin analogs were mainly bound to heavy proteins (mostly albumin) and to a small extent to low-density lipoproteins.

Simple ion chromatographic method for the determination of chlormequat residues in pears.

Current methods for quantitative determination of chlormequat residues in food crops are characterized by rather low recoveries and the need for derivatization (in case of gas chromatography, GC), or by high capital investment (in case of liquid chromatography-mass spectrometry, LC-MS). We propose a cation-exchange chromatography method for the analysis of chlormequat in pears. The method is based on extraction of the target compound with 40 mM HCl, followed by centrifugation and filtration. The filtrate is directly injected into an ion chromatograph equipped with a commercially available cation-exchange column and a suppressed conductivity detection system. While the limit of detection (LOD) (0.5 mg/kg) may not be small enough to allow dietary analysis, the method meets all validation requirements and is an alternative for the existing GC and LC-MS methods in quality control.

Cytotoxic activity of di-n-butyltin(IV)(X-A-B-Y) compounds related to salicylic acid against human tumour cells.

A series of di-n-butyltin(IV) compounds of the type di-n-butyltin(IV)(X-A-B-Y) with the (X-A-B-Y) being related to salicylic acid was tested in vitro for cytotoxic activity against tumour cells of the human A 204, MCF-7, T24, WiDr and IgR-37 cell lines using the PIT method. In each cell line the di-n-butyltin(IV) compounds displayed similar activity. As compared to cisplatin, the compounds were 4- to 10-fold more effective against A 204, MCF-7 and T24 tumour cells and slightly more effective against WiDr cells. The effectiveness against IgR-37 cells was similar for cisplatin and di-n-butyltin (IV) compounds.

Synthesis, characterization and in vitro antitumor activity of di- and triorganotin derivatives of polyoxa- and biologically relevant carboxylic acids.

An overview of the development of anti-tumor organotin derivatives, sometimes as active in vitro as doxorubicin, is presented and discussed. Solubility in water is an important issue, dominating the in vivo testing of compounds with promising in vitro properties. Several water-soluble organotin compounds gave the best in vitro activities. Novel, useful organotin anti-tumor compounds should be designed toward improved water solubility.

A comparative study of the photosensitizing characteristics of some cyanine dyes.

The present work has been carried out to explore the potential application of cyanines in photodynamic therapy. After photosensitization, the in vitro cytotoxic and antiproliferative activity on HeLa cells of a total of 35 cyanines belonging to several chemical subgroups is explored. Most of these cyanines have never been used before in similar experimental work. From a first set of experiments, it is found that none of the krypto-, oxa- and imidacyanines is photobiologically active on HeLa cells. Conversely, five thiacyanines (Thiacl-5), one rhodacyanine (Rhodac) and four indocyanines (Indoc2, Indoc4, Indoc5, Indoc7) show photodependent cytotoxicity or antiproliferative effects. A more detailed study shows that out of the ten selected compounds, eight cyanines feature significant photodependent cytotoxic and antiproliferative effects. All possess maximum absorption ranges between 545 and 824 nm. In particular, Rhodac, a tetramethinemeromonomethine rhodacyanine dye with an absorption maximum of 655 nm (ethanol) and a molar absorption coefficient epsilon = 108000 shows very promising photo-dependent biological activity. In general, the measured singlet oxygen quantum yield of the selected cyanines is low (< 0.08) and does not correlate with the degree of photosensitization. Furthermore, the present study shows that cyanines with a partition coefficient close to 1.5 accumulate to the highest extent in HeLa cells, while the more hydrophobic compounds (e.g., indocyanines) concentrate less intracellularly.